Novel lapacho compounds and methods of use thereof

ABSTRACT

The invention provides new synthetic lapacho derivatives as well as methods of use thereof. These compounds can be used in pharmaceutical compositions for the treatment or prevention of cell proliferation disorders. These compounds can also be used in the treatment or prevention of psoriasis or cancer or precancerous conditions.

BACKGROUND OF THE INVENTION

Lapacho (“pau d'arco”, “ipê-roxo”, “taheebo”) is a commercial naturalproduct obtained from the bark of Tabebuia trees, and in particular fromT. impetiginosa (Martius ex DC.) Standley (3inoniaceae), which are foundin the rainforests throughout Central and South America. Lapacho hasbeen used as a folk medicine for many years, in particular for thetreatment of cancer (Hartwell, J. L., Lloydia, 31, 71-170, 1968) anddisorders of the immune system, including psoriasis (Jones, I, PauD'arco: Immune Power from the Rain Forest; Healing Arts Press;Rochester, Vt., 1995).

The occurrence of naphthoquinones in various members of the genusTabebuia has been widely reported (Burnett, A. R., et. al., J. Chem.Soc., C, 2100-2104, 1967; Rao, M. M, et. al, J. Nat. Prod., 45, 600-604,1982; Girard, M., et. al., J. Nat. Prod., 51, 1023-1024, 1988; and Diaz,F., et. al., J. Nat. Prod., 59, 423424, 1996). The best known of thesecompounds are lapachol, alpha-lapachone (α-lapachone) and beta-lapachone(β-lapachone), which have the following chemical structures:

Although all three of these compounds have been reported to haveantiproliferative activity, β-lapachone, in particular, has demonstratedsignificant antineoplastic activity against a wide spectrum of humancancer cell lines at concentrations typically in the range of 1-10 μM(IC₅₀). For example, the cytotoxicity of β-lapachone has beendemonstrated in transformed cell lines derived from patients withpromyelocytic leukemia (Planchon et al., Cancer Res., 55 (1996) 3706),prostate ( Li, C J., et al., Cancer Res., 55 (1995) 3712), malignantglioma (Weller, M. et al., Int. J. Cancer, 73 (1997) 707), hepatoma(Lai, C. C., et al., Histol Histopathol, 13 (1998) 8), colon (Huang, L.,et al., Mol Med, 5, (1999) 711), breast (Wuertzberger, S. M., et al.,Cancer Res., 58 (1998) 1876), ovarian (Li, C. J. et al., Proc. Natl.Acad. Sci. USA, 96(23) (1999) 13369-74), pancreatic (Li, Y., et al., MolMed, 6 (2000) 1008; Li, Y. Z., Mol Med, 5 (1999) 232), and multiplemyeloma cell lines, including drug-resistant lines (Li, Y., Mol Med, 6(2000) 1008). No cytotoxic effects were observed on normal fresh orproliferating human PBMC (Li, Y., Mol Med, 6 (2000) 1008).

Other lapacho-derived compounds have been shown to haveantiproliferative activity. Eight compounds, representing the mostcommon constituents of the inner bark of T. impetiginosa and includinglapachol, α-lapachone and β-lapachone, were evaluated forantiproliferative and cytotoxic activity in the nontransformed humankeratinocyte cell line HaCaT, a model for the highly proliferativeepidermis characteristic of psoriasis (Müller, K., et al., J. Nat. Prod.62 (1999) 1134-1136). While lapachol and α-lapachone were relativelyinactive in this model, β-lapachone and several naphtho[2,3-b]furandiones displayed inhibition of keratinocyte growth comparable to theantipsoriatic drug anthralin. These findings encourage the design andsynthesis of new lapacho compounds and their evaluation forantiproliferative activity in a variety of biological systems.

SUMMARY OF THE INVENTION

The present invention provides new synthetic lapacho derivatives ofFormula I:

wherein X is O or S; and R is straight-chained or branched C₁-C₆ alkyl,aryl, substituted aryl (substituted, for example, with: hydroxyl,alkoxy, C₁-C₆ alkyl, nitro, halogen carboxyl, carboxyalkyl), orstraight-chained or branched alkylaryl, or a pharmaceutically acceptablesalt thereof; wherein 1) R is not methyl; 2) where X is O, R is notbromomethyl, unsubstituted phenyl, or phenyl substituted at the4-position with methyl, chloro, ethenyl, or 2′-chloroethyl; 3) where Xis S, R is not 2-carboxyphenyl. In preferred embodiments, R is an arylgroup; in more preferred embodiments, the aryl group is a phenyl groupsubstituted with one or two hydroxyl or alkyloxy groups (preferablyalkoxy groups); in still more preferred embodiments, the phenyl group issubstituted with one or two methoxy groups; more preferably, the phenylgroup is a 3,4-dimethoxyphenyl group.

The present invention also provides new synthetic lapacho derivatives ofFormula II:

wherein X is O or S; and R is straight-chained or branched C₁-C₆ alkyl,aryl, substituted aryl (substituted, for example, with: hydroxyl,alkoxy, C₁-C₆ alkyl, nitro, halogen carboxyl, carboxyalkyl), orstraight-chained or branched alkylaryl, or a pharmaceutically acceptablesalt thereof; wherein R is not methyl. In preferred embodiments, R is anaryl group; in more preferred embodiments, the aryl group is a phenylgroup substituted with one or two hydroxyl or alkyloxy groups; in stillmore preferred embodiments, the phenyl group is substituted with one ortwo methoxy groups; more preferably, the phenyl group is a3,4-dimethoxyphenyl group.

The present invention also concerns new synthetic lapacho analogs ofFormula III:

wherein X is O or S; R₁ is independently at each incidence hydroxyl,alkoxyl, C₁-C₆ alkyl, nitro, halogen, carboxyl or carboxyalkyl; R₂ ishydrogen or —C(O)—R₃, R₃ is straight-chained or branched C₁-C₆ alkyl,aryl, substituted aryl (substituted, for example, with: hydroxyl,alkoxy, C₁-C₆ alkyl, nitro, halogen, carboxyl, carboxyalkyl), orstraight-chained or branched alkylaryl; and n is 0, 1 or 2; or apharmaceutically acceptable salt thereof; wherein 1) where X is O, R₂ isnot H; 2) where X is O, and R₂ is —C(O)—R₃, and R₃ is methyl, then R₁ isnot hydroxyl or methoxy; and 3) where X is S and R₂ is H, then n is 1and R₁ is selected from —OH and —OC(O)-alkyl(C₁-C₆); and 4) where X is Sand R₂ is —C(O)—R₃, and R₃ is methyl, then R₁ does not represent a7-acetyl group.

Preferred compounds of Formula I are those in which X is S and R is arylor substituted aryl.

Preferred compounds of Formula II are those in which X is O or S and Ris alkyl, aryl or mono- or di-substituted aryl.

Preferred compounds of Formula III are those in which X is S, R₁ ishydroxyl or —OC(O)-alkyl(C₁-C₆), R₂ is hydrogen, and n is 1; still morepreferably, R₁ is 5-hydroxyl or 5—OC(O)-methyl.

The present invention also provides pharmaceutical formulationscomprising a compound of Formula I, II or III in combination with atleast one pharmaceutically acceptable excipient or carrier.

The present invention also provides a method for the treatment of cellproliferative disorders in mammals comprising administering to a mammalin need of such treatment an effective amount of a compound of FormulaI, II or III. The invention further provides the use of a compound ofFormula I, II or III for the preparation of a medicament useful for thetreatment of a cell proliferative disorder.

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. Although methods and materialssimilar or equivalent to those described herein can be used in thepractice or testing of the present invention, suitable methods andmaterials are described below. All publications, patent applications,patents, and other references mentioned herein are incorporated byreference in their entirety. In the case of conflict, the presentspecification, including definitions, will control. In addition, thematerials, methods, and examples are illustrative only and are notintended to be limiting.

Other features and advantages of the invention will be apparent from thefollowing detailed description and claims.

DETAILED DESCRIPTION OF THE INVENTION

The present invention provides new synthetic lapacho derivatives ofFormula I:

wherein X is O or S; and R is straight-chained or branched C₁-C₆ alkyl,aryl, substituted aryl (substituted, for example, with from one to fourof the following moieties: hydroxyl, alkoxy, C₁-C₆ alkyl, nitro,carboxyl, carboxyalkyl), or straight-chained or branched alkylaryl, or apharmaceutically acceptable salt thereof; wherein 1) R is not methyl; 2)where X is O, R is not bromomethyl, unsubstituted phenyl, or phenylsubstituted at the 4-position with methyl, chloro, ethenyl, or2′-chloroethyl; 3) where X is S, R is not 2-carboxyphenyl. In preferredembodiments, R is an aryl group; in more preferred embodiments, the arylgroup is a phenyl group substituted with one or two hydroxyl or alkoxygroups (preferably alkoxy groups); in still more preferred embodiments,the phenyl group is substituted with one or two methoxy groups; morepreferably, the phenyl group is a 3,4-dimethoxyphenyl group.

Preferred compounds of Formula I are those in which X is S and R is arylor substituted aryl. For example, aryl can have 1, 2, 3, or 4substituents, which can be the same or different. Preferably, the arylgroup has 1, 2, 3, or 4 substituents independently selected fromhydroxyl, alkoxy, alkyl, nitro, halogen, carboxyl or carboxyalkyl.

The present invention also provides new synthetic lapacho derivatives ofFormula II:

wherein X is O or S; and R is straight-chained or branched C₁-C₆ alkyl,aryl, substituted aryl (substituted, for example, with from one to fourof the following moieties: hydroxyl, alkoxy, C₁-C₆ alkyl, nitro,carboxyl, carboxyalkyl), or straight-chained or branched allylaryl, or apharmaceutically acceptable salt thereof; wherein R is not methyl. Inpreferred embodiments, R is an aryl group; in more preferredembodiments, the aryl group is a phenyl group substituted with one ortwo hydroxyl or alkyloxy groups; in still more preferred embodiments,the phenyl group is substituted with one or two methoxy groups; morepreferably, the phenyl group is a 3,4-dimethoxyphenyl group.

Preferred compounds of Formula II are those in which X is O or S and Ris alkyl, aryl or mono- or di-substituted aryl. For example, aryl canhave 1, 2, 3, or 4 substituents, which can be the same or differentPreferably, the aryl group has 1, 2, 3, or 4 substituents independentlyselected from hydroxyl, alkoxy, alkyl, nitro, halogen, carboxyl orcarboxyalkyl.

The present invention also concerns new synthetic lapacho analogs ofFormula III:

wherein X is O or S; R₁ is independently at each incidence hydroxyl,alkoxyl, C₁-C₆ alkyl, nitro, halogen, carboxyl or carboxyalkyl; R₂ ishydrogen or —C(O)—R₃, R₃ is straight-chain or branched C₁-C₆ alkyl,aryl, substituted aryl (substituted, for example, with from one to fourof the following moieties: hydroxyl, alkoxy, C₁-C₆ alkyl, nitro,carboxyl, carboxyalkyl), or straight-chained or branched alkylaryl; andn is 0, 1 or 2; or a pharmaceutically acceptable salt thereof;wherein 1) where X is O, R₂ is not H; 2) where X is O, and R₂ is—C(O)—R₃, and R₃ is methyl, then R₁ is not hydroxyl or methoxy; and 3)where X is S and R₂ is H, then n is 1 and R₁ is selected from —OH and—OC(O)-alkyl(C₁-C₆); and 4) where X is S and R₂ is —C(O)—R₃, and R₃ ismethyl, then R₁ does not represent a 7-acetyl group.

Preferred compounds of Formula III are those in which X is O, R₁ ishydroxyl or —OC(O)-alkyl(C₁-C₆), R₂ is hydrogen, and n is 1; still morepreferably, R₁ is 5-hydroxyl or 5—OC(O)-methyl.

Other preferred compounds of Formula III are those in which X is O or S,R₂ is hydrogen or —C(O)—R₃. In embodiments where R₂ is —C(O)—R₃, R₃ ispreferably an aryl group. For example, aryl can have 1, 2, 3, or 4substituents, which can be the same or different. Preferably, the arylgroup has 1, 2, 3, or 4 substituents independently selected fromhydroxyl, alkoxy, alkyl, nitro, halogen, carboxyl or carboxyalkyl.

In more preferred embodiments, the aryl group is a substituted orunsubstituted phenyl group. In certain preferred compounds of FormulaIII where R₂ is —C(O)—R₃, R₃ is a phenyl group substituted with one ortwo hydroxyl or alkyloxy groups; in still more preferred embodiments,the phenyl group is substituted with one or two methoxy groups; morepreferably, the phenyl group is a 3,4-dimethoxyphenyl group, whereinwhen n is 0, R₃ is not methyl.

Certain preferred compounds of the invention are shown in Tables 1-4.

The term “alkyl” refers to radicals containing carbon and hydrogen,without unsaturation. Alkyl radicals can be straight or branched.Exemplary alkyl radicals include, without limitation, methyl, ethyl,propyl, isopropyl, hexyl, t-butyl, sec-butyl and the like. A C₁-C₆ alkylgroup is an alkyl group having from one to six carbon atoms in thestraight or branched alkyl backbone. Alkyl groups optionally can besubstituted with one or more moieties such as hydroxyl group,carboxylate, oxo, halogen, thiol, cyano, nitro, amino, acylamino, C₁-C₆alkylthio, arylthio, C₁-C₆ alkyl, C₁-C₆ alkoxy, aryloxy,alkylcarbonyloxy, arylcarbonyloxy, C₃-C₆ cycloalkyl, C₃-C₆cycloalkyloxy, C₂-C₆ alkenyl, C₂-C₆ alkynyl, aryl, aminocarbonyl, C₁- C₆alkylcarbonyl, C₃-C₆ cycloalkylcarbonyl, heterocyclylcarbonyl,arylcarbonyl, aryloxycarbonyl, C₁-C₆ alkoxycarbonyl, C₃-C₆cycloalkyloxycarbonyl, heterocyclyloxycarbonyl, C₁-C₆ alkylsulfonyl,arylsulfonyl, a heterocyclyl group, and the like.

Alkyl radicals can be cyclic. A “cycloalkyl” group refers to a cyclicalkyl group which has a ring having from three to six carbon atoms inthe ring portion. A cycloalkyl group may be substituted with one ormoieties as described for alkyl groups.

As used herein, the term “aryl” refers to an aromatic carbocyclic orheteroaromatic moiety, having one, two, or three rings. An aryl groupmay be carbocyclic or may optionally contain from 1-4 heteroatoms (suchas nitrogen, sulfur, or oxygen) in the aromatic ring. Exemplary arylgroups include, without limitation, phenyl, naphthyl, pyridyl,pyrimidyl, triazinyl, quinazolinyl, thiazolyl, benzothiophenyl, furanyl,imidazolyl, thiophenyl and the like. An aryl group optionally can besubstituted with one or more substituents such as hydroxyl group,halogen, thiol, cyano, nitro, amino, acylamino, C₁-C₆ alkylthio,arylthio, C₁-C₆ alkyl, C₁-C₆ alkoxy, aryloxy, alkylcarbonyloxy,arylcarbonyloxy, C₃-C₆ cycloalkyl, C₃-C₆ cycloalkyloxy, C₂-C₆ alkenyl,C₂-C₆ alkynyl, aryl, carboxylate, aminocarbonyl, C₁-C₆ alkylcarbonyl,C₃-C₆ cycloalkylcarbonyl, heterocyclylcarbonyl, arylcarbonyl,aryloxycarbonyl, C₁-C₆ alkoxycarbonyl, C₃-C₆ cycloalkyloxycarbonyl,heterocyclyloxycarbonyl, aryloxycarbonyl, C₁-C₆ alkoxycarbonyl, C₁-C₆alkylsulfonyl, arylsulfonyl, a heterocyclyl group, and the like.

The term “heterocyclyl” or “heterocycle” refers to a stable non-aromatic3-7 membered monocyclic heterocyclic ring or 7-11 membered bicyclicheterocyclic ring which is either saturated or unsaturated, and may befused, spiro or bridged to form additional rings. Each heterocycleconsists of one or more carbon atoms and from one to four heteroatomsselected from the group consisting of nitrogen, oxygen and sulfur. Aheterocyclyl radical may be attached at any endocyclic atom whichresults in the creation of a stable structure. Preferred heterocyclesinclude 3-7 membered monocyclic heterocycles (more preferably5-7-membered monocyclic heterocycles) such as (without limitation)piperidinyl, pyranyl, piperazinyl, morpholinyl, thiamorpholinyl, andtetrahydrofuranyl.

In general, structures depicted herein are meant to include allstereochemical forms of the structure; i.e., the R and S configurationsfor each asymmetric center, unless a particular stereochemistry isspecifically indicated. Therefore, single stereochemical isomers (i.e.,substantially pure enantiomers and diasteromers) as well as enantiomericand diastereomeric mixtures, such as racemic mixtures, of the presentcompounds are within the scope of the invention. Furthermore, allgeometric isomers, such as E- and Z-configurations at a double bond, arewithin the scope of the invention unless otherwise stated. Certaincompounds of this invention may exist in tautomeric forms. All suchtautomeric forms of the compounds are considered to be within the scopeof this invention unless otherwise stated.

The present invention also provides pharmaceutical formulationscomprising a compound of Formula I, II or III in combination with atleast one pharmaceutically acceptable excipient or carrier. As usedherein, “pharmaceutically acceptable excipient” or “pharmaceuticallyacceptable carrier” is intended to include any and all solvents,dispersion media, coatings, antibacterial and antifungal agents,isotonic and absorption delaying agents, and the like, compatible withpharmaceutical administration. Suitable carriers are described in“Remington: The Science and Practice of Pharmacy, Twentieth Edition,”Lippincott Williams & Wilkins, Philadelphia, Pa., which is incorporatedherein by reference. Preferred examples of such carriers or diluentsinclude, but are not limited to, water, saline, Ringer's solutions,dextrose solution, and 5% human serum albumin. Liposomes and non-aqueousvehicles such as fixed oils may also be used. The use of such media andagents for pharmaceutically active substances is well known in the art.Except insofar as any conventional media or agent is incompatible withthe active compound, use thereof in the compositions is contemplated.Supplementary active compounds can also be incorporated into thecompositions.

A compound of Formula I, II, or III is administered in a suitable dosageform prepared by combining a therapeutically effective amount (e.g., anefficacious level sufficient to achieve the desired therapeutic effectthrough inhibition of tumor growth, killing of tumor cells, etc.) of acompound of Formula I, II, or III (as an active ingredient) withstandard pharmaceutical carriers or diluents according to conventionalprocedures (i.e., by producing a pharmaceutical composition of theinvention). These procedures may involve mixing, granulating, andcompressing or dissolving the ingredients as appropriate to attain thedesired preparation.

Preferred pharmaceutically acceptable carriers include solid carrierssuch as lactose, terra alba, sucrose, talc, gelatin, agar, pectin,acacia, magnesium stearate, stearic acid and the like. Exemplary liquidcarriers include syrup, peanut oil, olive oil, water and the like.Similarly, the carrier or diluent may include time-delay material knownin the art, such as glyceryl monostearate or glyceryl distearate, aloneor with a wax, ethylcellulose, hydroxypropylmethylcellulose,methylmethacrylate or the like. Other fillers, excipients, flavorants,and other additives such as are known in the art may also be included ina pharmaceutical composition according to this invention.

The pharmaceutical compositions containing active compounds of thepresent invention may be manufactured in a manner that is generallyknown, e.g., by means of conventional mixing, dissolving, granulating,dragee-making, levigating, emulsifying, encapsulating, entrapping, orlyophilizing processes. Pharmaceutical compositions may be formulated ina conventional manner using one or more physiologically acceptablecarriers comprising excipients and/or auxiliaries which facilitateprocessing of the active compounds into preparations that can be usedpharmaceutically. Of course, the appropriate formulation is dependentupon the route of administration chosen.

A compound or pharmaceutical composition of the invention can beadministered to a subject in many of the well-known methods currentlyused for chemotherapeutic treatment For example, for treatment ofcancers, a compound of the invention may be injected directly intotumors, injected into the blood stream or body cavities or taken orallyor applied through the skin with patches. For treatment of psoriaticconditions, systemic administration (e.g., oral administration), ortopical administration to affected areas of the skin, are preferredroutes of administration. The dose chosen should be sufficient toconstitute effective treatment but not so high as to cause unacceptableside effects. The state of the disease condition (e.g., cancer,psoriasis, and the like) and the health of the patient should preferablybe closely monitored during and for a reasonable period after treatment.

The present invention also provides a method for the treatment of cellproliferative disorders in mammals comprising administering to a mammalan effective amount of a compound of Formula I, II or III. The mammal ispreferably a mammal in need of such treatment. The mammal can be e.g.,any mammal, e.g., a human, a primate, mouse, rat, dog, cat, cow, horse,pig. In a preferred embodiment, the mammal is a human. The inventionfurther provides the use of a compound of Formula I, II or III for thepreparation of a medicament useful for the treatment of a cellproliferative disorder. The compounds of the invention are preferablyadministered in the form of pharmaceutical compositions, e.g., asdescribed herein.

As used herein, the term “cell proliferative disorder” refers toconditions in which the unregulated and/or abnormal growth of cells canlead to the development of an unwanted condition or disease, which canbe cancerous or non-cancerous, for example a psoriatic condition. Asused herein, the term “psoriatic condition” refers to disordersinvolving keratinocyte hyperproliferation, inflammatory cellinfiltration, and cytokine alteration.

In addition to psoriatic conditions, the types of proliferative diseaseswhich may be treated using the compositions of the present invention areepidermic and dermoid cysts, lipomas, adenomas, capillary and cutaneoushemangiomas, lymphangiomas, nevi lesions, teratomas, nephromas,myofibromatosis, osteoplastic tumors, and other dysplastic masses andthe like.

The invention will be further described in the following examples, whichdo not limit the scope of the invention described in the claims.

EXAMPLES Example 1. Synthesis of Synthetic Lapcho Analogs

Compounds of the invention can be prepared in a variety of ways, some ofwhich are known in the art. In general, the compounds of the presentinvention can be prepared from commercially available startingmaterials, compounds known in the literature, or from readily-preparedintermediates, by employing standard synthetic methods and proceduresknown to those skilled in the art, or which will be apparent to theskilled artisan in light of the teachings herein. Standard syntheticmethods and procedures for the preparation of organic molecules andfunctional group transformations and manipulations can be obtained fromthe relevant scientific literature or from standard textbooks in thefield. Although not limited to any one or several sources, classic textssuch as Smith, M. B.; March, J. March's Advanced Organic Chemistry:Reactions, Mechanisms, and Structure, 5^(th) ed.; John Wiley & Sons: NewYork, 2001; and Greene, T. W.; Wuts, P. G. M. Protective Groups inOrganic Synthesis, 3^(rd).; John Wiley & Sons: New York, 1999 are usefuland recognized reference textbooks of organic synthesis known to thosein the art. The following descriptions of synthetic methods are designedto illustrate, but not limit, general procedures for the preparation ofcompounds of the invention.

Melting points were determined with a Reichert Thermovar melting pointapparatus and are uncorrected. Chromatography refers to columnchromatography on silica gel (E. Merck, 70-230 mesh) using CH₂Cl₂ aseluant, unless otherwise stated. ¹H NMR spectra were recorded with aVarian EM 390 (90 MHz) or a Bruker Spectrospin WM 250 spectrometer (250MHz), using tetramethylsilane as an internal standard. Fourier-transformIR spectra (KBr) were recorded on a Nicolet 510M FTIR spectrometer. UVspectra were recorded on a Kontron 810 spectrometer. Mass spectra (EI)were obtained on a Varian MAT 112S spectrometer (70 eV). ElementalAnalysis were within ±0.4% of calculated values.

Example 2. Analogs of Formulae I and II

One process that can be used for the preparation of the most preferredcompounds of Formula I and II is shown in Scheme 1. Introduction of the2-acyl functionality onto the naphtho[2,3-b]thiophene andnaphtho[2,3-b]furan nuclei was achieved by metalation withsec-butyllithium in the presence of tetramethylethylenediamine, wheresubstitution occurs exclusively in the 2-position. The reaction ofnaphtho[2,3-b]thiophene- and naphtho[2,3-b]furan-2-yl-lithium with theappropriate aldehydes gave the secondary alcohols 62-66 and 73-75,respectively. The desired acyl group was obtained by oxidation of thealcohol group with activated manganese(IV) oxide in methylene chloride.Oxidation of the 2-acyl analogues with chromium trioxide in glacialacetic acid provided the corresponding naphtho[2,3-b]thiophene- andnaphtho[2,3-b]furan4,9-diones 62b-66b and 6, 74b, 75b, respectively. Thephenolic analogues 62c and 65c were obtained by ether cleavage of thecorresponding methyl ethers 62b and 65b with boron tribromide inmethylene chloride.

Reagents: (a) sec-BuLi, tetramethylethylenediamine, ether, −78°; (b)MnO₂, CH₂Cl₂; (c) CrO₃, HOAc; (d) BBr₃, CH₂Cl₂. R and X are defined inTables 1 and 2.

Compounds of Scheme 1

General Procedure for the Preparation of(R,S)-Naphtho[2,3-b]thiophen-2-yl-alkanols.(R,S)-(4-Methoxyphenyl)-naphtho[2,3-b]thiophen-2-yl-methanol (62). To asolution of naphtho[2,3-b]thiophene (0.92 g, 4.99 mmol) in absolute Et₂O(80 mL) and tetramethylethylenediamine (0.08 mL, 0.8 mmol) was addedsec-butyllithium (4.40 mL of a 1.3 M solution in hexane, 5.72 mmol) at−78° C. under N₂. Then the solution was stirred at −78° C. for 1 h. Dry4-methoxybenzaldehyde (0.73 mL, 6.0 mmol), freshly distilled, was addedat −78° C., and the solution was allowed to warm to room temperaturewithin 12 h. Then it was treated with a solution of half-saturated NH₄Cl(400 mL), the organic layer was washed with water (400 mL), dried overNa₂SO₄, and evaporated. The residue was purified by chromatography andrecrystallized from CH₂Cl₂/hexane to afford white crystals: 72% yield;mp 164-165° C.; FTIR 3377 (OH), 1611 cm⁻¹; ¹H NMR (CDCl₃) δ 8.25-6.90(m, 11H), 6.90 (d, 1H, ³J=4.01 Hz), 3.82 (s, 3H), 2.47 (d, 1H, ³J=4.01Hz, exchangeable); MS, m/z 320 (66, M³⁰ ), 135 (100). Anal. (C₂₀H₁₆O₂S)C, H.

(R,S)-Naphtho[2,3-b]thiophen-2-yl-ethanol (63) was obtained fromnaphtho[2,3-b]thiophene (1.00 g, 5.43 mmol) and acetaldehyde (0.37 mL,6.52 mmol) as described for 62 to afford white needles: 44% yield; mp182-184° C.; FTIR 3319 (OH) cm¹; ¹H NMR (CDCl₃) δ8.23-7.43 (m, 6H), 7.25(s, 1H), 5.21 (qu, 1H, ³J=6.42 Hz), 1.82 (d, 1H, ³J=6.42 Hz,exchangeable), 1.69 (s, 3H). Anal. (C₁₄H₁₂OS) C, H.

(R,S)-Naphtho[2,3-b]thiophen-2-yl-phenylmethanol (64) was obtained fromnaphtho[2,3-b]thiophene (0.75 g, 4.07 mmol) and benzaldehyde (0.49 mL,4.88 mmol) as described for 62 to afford white needles: 67% yield; mp142-145° C.; FTIR 3319 (OH) cm⁻¹; ¹H NMR (CDCl₃) δ8.56-7.16 (m, 12H),6.13 (d, 1H, ³J=3.59 Hz), 2.58 (d, 1H, ³J=3.59 Hz, exchangeable). Anal.(C₁₉H₁₄OS) C, H.

(R,S)-(3,4-Dimethoxyphenyl)-naphtho[2,3-b]thiophen-2-yl-methanol (65)was obtained from naphtho[2,3-b]thiophene (1.00 g, 5.43 mmol) and3,4-dimethoxybenzaldehyde (1.08 g, 6.51 mmol) as described for 62 toafford white crystals: 65% yield; mp 175-176° C.; FTIR 3481 (OH), 1594cm⁻¹; ¹H NMR (CDCl₃) δ8.43-6.93 (m, 10H), 6.40 (d, 1H,³J=3.00 Hz,exchangeable), 6.00 (d, 1H, ³J=3.00 Hz), 3.81 (s, 3H), 3.80 (s, 3H).Anal. (C₂₁H₁₈O₃S) C, H.

(R,S)-Naphtho[2,3b]thiophen-2-yl-(4-nitrophenyl)methanol (66) wasobtained from naphtho[2,3-b]thiophene (1.00 g, 5.43 mmol) and4nitrobenzaldehyde (1.07 g, 7.06 mmol) as described for 62 to affordlight-yellow crystals: 55% yield; mp 239-240° C.; FTIR 3548 (OH), 1596cm⁻¹; ¹H NMR (DMSO-d₆) δ8.67-7.43 (m, 11H), 6.84 (d, 1I, ³J=4.31 Hz,exchangeable), 6.27 (d, 1H, ³J=4.31 Hz). Anal. (C₁₉H₁₃NO₃S) C, H.

General Procedure for the Preparation ofNaphtho[2,3-b]thiophen-2-yl-akanones.(4-Methoxyphenyl)-naphtho[2,3-b]thiophen-2-yl-methanone (62a). To asolution of 62 (1.00 g, 3.12 mmol) in CH₂Cl₂ (100 mL) was addedactivated MnO₂ (2.61 g, 30 mmol), and the mixture was stirred for 1.5 h,until the oxidation was completed (TLC control). The suspension wasfiltered, and the residue was washed with CH₂Cl₂ (3×200 mL). Thesolution was treated with hexane (500 mL), then concentrated, and theproduct was crystallized at −18° C. to afford lemon crystals: 81% yield;mp 211-212° C.; FTIR 1622 (CO), 1603 cm⁻¹; ¹H NMR (CDCl₃) δ8.43-7.01 (m,11H), 3.92 (s, 3H); MS nmz 318 (100, M³⁰ ). Anal. (C₂₀H₁₄O₂S) C, R

Naphtho[2,3-b]thiophen-2-yl-ethanone (63a) was obtained from 63 (0.35 g,1.53 mmol) as described for 62a, but it was stirred for 24 h to affordgreenish-yellow needles: 84% yield; mp 224-225° C.; FTIR 1663 (CO) cm⁻¹;¹H NMR (DMSO-d₆) 68.67-7.51 (m, 7H), 2.71 (s, 3H). Anal. (C₁₄H₁₀OS) C,H.

Naphtho[2,3-b]thiophen-2-yl-phenylmethanone (64a) was obtained from 64(0.35 g, 1.21 mmol) as described for 62a to afford yellow crystals: 96%yield; mp 164-165° C.; FTIR 1630 (CO), 1595 cm⁻¹; ¹H NMR (CDCl₃)δ8.42-7.45 (m, 12H). Anal. (C₁₉H₁₂OS) C, H.

(3,4-Dimethoxyphenyl)-naphtho[2,3-b]thiophen-2-yl-methanone (65a) wasobtained from 65 (0.91 g, 2.60 mmol) as described for 62a to affordlemon crystals: 87% yield; mp 175-176° C.; FTIR 1630 (CO), 1596 cm⁻¹; ¹HNMR (CDCl₃) 88.44-6.97 (m, 10H), 4.00 (s, 3H), 3.98 (s, 3H). Anal.(C₂₁H₁₆O₃S) C, H.

Naphtho[2,3-b]thiophen-2-yl-(4-nitrophenyl)methanone (66a) was obtainedfrom 66 (0.50 g, 1.49 mmol) as described for 62a to afford orangecrystals: 97% yield; mp 251-252° C.; FTIR 1628 (CO), 1600 cm⁻¹; ¹H NMR(CDCl₃) δ8.78-7.48 (m, 11H). Anal. (C₁₉H₁₁NO₃S) C, H.

General Procedure for the Oxidation of Naphtho[2,3-b]thiophenes toNaphtho[2,3-b]thiophene-4,9-diones.2-(4-Methoxybenzoyl)-naphtho[2,3-b]thiophene-4,9-dione (62b). To asolution of 62a (0.75 g, 2.35 mmol) in glacial acetic acid (50 mL) wasadded with stirring at room temperature, dropwise over 1 h, a solutionof CrO₃ (0.66 g, 6.6 mmol) in glacial acetic acid (10 mL) and water (10mL). The solution was stirred for an additional 30 min, then water (250mL) was added, the product was filtered by suction, and purified bychromatography. The combined fractions were treated with hexane, thenconcentrated, and the product was crystallized at −18° C. to affordlemon needles: 86% yield; mp 165-166° C.; FTIR 1667 (CO), 1600 cm⁻¹; ¹HNMR (CDCl₃) δ8.29-7.00 (m, 9H), 3.92 (s, 3M; MS m/z 348 (92, M³⁰ ), 135(100). Anal. (C₂₀H₁₂O₄S) C, H.

2-Acetyl-naphtho[2,3-b]thiophene-4,9-dione (63b) was obtained from 63a(0.18 g, 0.80 mmol) as described for 62b to afford yellow crystals: 64%yield; mp 261-262° C.; FTIR 1669 (CO), 1651 (CO), 1590 cm⁻¹; ¹H NMR(DMSO-d₆) δ8.44 (s, 1H), 8.21-7.92 (m, 4H), 2.71 (s, 3H). Anal.(C₁₄H₈O₃S) C, H.

2-Benzoyl-naphtho[2,3-b]thiophene-4,9-dione (64b) was obtained from 64a(0.35 g, 1.21 mmol) as described for 62b to afford yellow crystals: 70%yield; mp 158-159° C.; FTIR 1667 (CO), 1652 (CO), 1634 (CO), 1594 cm⁻¹;¹H NMR (CDCl₃) δ8.30-7.53 (m, 10H). Anal. (C₁₉H₁₀O₃S) C, H.

2-(3,4-Dimethoxybenzoyl)-naphtho[2,3-b]thiophene-4,9-dione (65b) wasobtained from 65a (0.57 g, 1.63 mmol) as described for 62b to affordorange-yellow needles: 48% yield; mp 212-218° C.; FTIR 1669 (CO), 1629(CO), 1593 cm⁻¹; ¹H NMR (DMSO-d₆) δ8.19-7.18 (m, 8H), 3.91 (s, 3H), 3.86(s, 3H). Anal. (C₂₁H₁₄O₅S) C, H.

2-(4-Nitrobenzoyl)-naphtho[2,3-b]thiophene-4,9-dione (66b) was obtainedfrom 66a (0.30 g, 0.90 mmol) as described for 62b to afford lemonneedles: 76% yield; mp 270° C.; FTIR 1671 (CO), 1640 (CO) cm⁻¹; ¹H NMR(CDCl₃) δ8.46-7.81 (m, 9H). Anal. (C₁₉H₉NO₅S) C, H.

General Procedure for the Cleavage of Methyl Ethers.2-(4-Hydroxybenzoyl)-naphtho[2,3-b]thiophene-4,9-dione (62c). To asolution of 62b (0.25 g, 0.72 mmol) in dry CH₂Cl₂ (50 mL) was added BBr₃(0.70 mL, 7.18 mmol) at room temperature under N₂, and the solution wasstirred at room temperature for 120 h. Then 2 N HCl (100 mL) was added,the organic layer was extracted with 2 N NaOH (3×100 mL), the combinedaqueous layer was acidified with conc. HCl, and the product wasdissolved in ethyl acetate. The organic layer was washed with asaturated solution of NaCl, then concentrated, and the product wascrystallized at −18° C. to afford yellow-green crystals: 46% yield; mp264-265° C.; FTIR 3553 (OH), 1669 (CO), 1648 (CO) cm⁻¹; ¹H NMR (DMSO-d₆)δ10.66 (s, 1H, exchangeable), 8.19-6.96 (m, 9H). Anal. (C₁₉H₁₀O₄S) C, H.

2-(3,4-Dihydroxybenzoyl)-naphtho[2,3-b]thiophene-4,9-dione (65c) wasobtained from 65b (0.20 g, 0.53 mmol) as described for 62c.Recrystallization from toluenelethyl acetate afforded orange-yellowcrystals: 65% yield; mp 272-274° C. dec; FTIR 3448 (OH), 3309 (OH), 1671(CO), 1632 (CO) cm⁻¹; ¹H NMR (DMSO-d₆) δ10.0-9.0 (s, 2H, exchangeable),8.20-6.93 (m, 8H). Anal. (C₁₉H₁₀O₅S) C, H.

General Procedure for the Preparation of(R,S)-Naphtho[2,3-b]furan-2-yl-alkanols.(R,S)-Naphtho[2,3-b]furan-2-yl-ethanol (73). To a solution ofnaphtho[2,3-b]furan (0.50 g, 2.96 mmol) in absolute Et₂O (50 mL) andtetramethylethylenediamine (0.15 mL, 1.5 mmol) was addedsec-butyllithium (5.50 mL of a 1.3 M solution in hexane, 7.15 mmol) at−78° C. under N₂, and the solution was stirred at −78° C. for 4 h. Dryacetaldehyde (0.13 mL, 3.84 mmol), freshly distilled, was added at −78°C., and the solution was allowed to warm to room temperature within 12h. Then it was treated with a half-saturated solution of NH₄Cl (250 mL),the organic layer was washed with water (250 mL), dried over Na₂SO₄, andevaporated. The residue was purified by chromatography andrecrystallized from CH₂Cl₂/hexane to afford white needles: 65% yield; mp165-167° C.; FTIR 3313 (OH) cm⁻¹; ¹H NMR (CDCl₃) δ7.99-7.19 (m, 6H),6.72 (s, 1H), 5.6 (qu, 1H, ³J=6.6 Hz), 2.09 (s, 1H, exchangeable), 1.68(d, 3H, ³J=6.6 Hz). Anal. (C₁₄H₁₂O₂) C, H.

(R,S)-Naphtho[2,3-b]furan-2-yl-phenylmethanol (74) was obtained fromnaphtho[2,3-b]furan (0.32 g, 1.89 mmol) and benzaldehyde (0.25 mL, 2.46mmol) as described for 73. Recrystallization from hexane afforded whitecrystals: 42% yield; mp 100-101° C.; FTIR 3382 (OH) cm⁻¹; ¹H NMR (CDCl₃)δ7.96-7.32 (m, 11H), 6.65 (s, 1H), 5.98 (d, 1H, ³J=4.09 Hz), 2.58 (d,1H, ³J=4.09 Hz, exchangeable). Anal. (C₁₉H₁₄O₂) C, H.

General Procedure for the Preparation ofNaphtho[2,3-b]furan-2-yl-alkanones. Naphtho[2,3-b]furan-2-yl-ethanone(73a) was obtained from 73 (0.20 g, 0.94 mmol) as described for 62a, butit was stirred for 12 h to afford greenish needles: 81% yield; mp 193°C. dec (lit (Garuti et al. Farmaco Ed. Sci. 38: 527-532, 1983) 180° C.);FTIR 1679 (CO), 1632 cm⁻¹; ¹H NMR (CDCl₃) δ8.23-7.42 (m, 711), 2.67 (s,311). Anal. (C₁₄H₁₀O₂) C, H. Naphtho[2,3-b]furan-2-yl-phenylmethanone(74a) was obtained from 74 (0.18 g, 0.66 mmol) as described for 62.Recrystallization from hexane afforded yellow needles: 86% yield; mp140-142° C. (lit (Sen and Saxena, J. Indian Chem. Soc. 36: 283-284,1959) 101° C.); FTIR 1634 (CO) cm⁻¹; ¹H NMR (CDCl₃) δ8.25-7.43 (m, 12H).Anal. (C₁₉H₁₂O₂) C, H.

(3,4-Dimethoxyphenyl)-naphtho[2,3-b]furan-2-yl-methanone (75a) wasobtained from naphtho[2,3-b]furan (0.20 g, 1.18 mmol) and3,4-dimethoxybenzaldehyde (0.26 g, 1.54 mmol) as described for 73, andthe crude product was oxidized as described for 62a. Recrystallizationfrom hexane afforded light-yellow crystals: 61% yield; mp 155-157° C.;FTIR 1644 (CO) cm⁻¹; ¹H NMR (CDCl₃) δ8.25 (s, 1H), 8.04 (s, 1H),7.80-6.99 (m, 8H), 4.01 (s, 3H), 4.00 (s, 3H). Anal. (C₂₁H₁₆O₄) C, H.

General Procedure for the Preparafion of2-Acyl-naphtho[2,3-b]furan-4,9-diones.2-Acetyl-naphtho[2,3-b]furan4,9-dione (6) was obtained from 73a (0.11 g,0.80 mmol) as described for 62b. Recrystallization from CH₂Cl₂/hexaneafforded yellow crystals: 48% yield; mp 229-230° C. (lit (Lopez et al.J. Heterocycl Client. 21: 621-622, 1984) 222-224° C.); FTIR 1692 (CO),1674 (CO), 1582 cm⁻¹; ¹H NMR (CDCl₃) δ8.29-7.92 (m, 4H1), 7.61 (s, 1H),2.67 (s, 3H). Anal. (C₁₄H₈O₄) C, H.

2-Benzoyl-naphtho[2,3-b]furan-4,9-dione (74b) was obtained from 74a(0.05 g, 0.18 mmol) as described for 62b to afford yellow needles: 44%yield; mp 198-200° C.; FTIR 1674 (CO), 1659 (CO) cm⁻¹; ¹H NMR (CDCl₃)δ8.30-7.54 (m, 10H). Anal. (C₁₉H₁₀O₄) C, H.

2-(3,4-Dimethoxybenzoyl)-naphtho[2,3-b]furan:4,9-dione (75b) wasobtained from 75a (0.16g, 0.48 mmol) as described for 62b.Recrystallization from CH₂Cl₂/hexane afforded lemon crystals: 40% yield;mp 242-243° C.; FTIR 1676 (CO), 1638 (CO) cm⁻¹; ¹H NMR (CDCl₃)δ8.31-7.00 (m, 8H), 4.01 (s, 3H), 4.00 (s, 3H). Anal. (C₂₁H₁₄O₆) C, H.

Various compounds of formulae I and II are shown in Tables 1 and 2,below. TABLE 1 I

Compound X R 73a O Me 74a O Ph 75a O 3,4-(OMe)₂-Ph 63a S Me 64a S Ph 62aS 4-OMe-Ph 65a S 3,4-(OMe)₂-Ph 66a S 4-NO₂-Ph

TABLE 2 II

Compound X R 6 O Me 74b O Ph 75b O 3,4-(OMe)₂-Ph 63b S Me 64b S Ph 62b S4-OMe-Ph 65b S 3,4-(OMe)₂-Ph 62c S 4-OH-Ph 65c S 3,4-(OH)₂-Ph 66b S4-NO₂-Ph

Example 3 Analogs of Formula III

The process used for the preparation of most preferred compounds ofFormula III is shown in Scheme 2. An aluminum chloride catalyzedFriedel-Crafts acylation of thiophene with 3-hydroxyphthalic anhydridein methylene chloride afforded exclusively2-hydroxy-6-(2-thenoyl)-benzoic acid (77), which was formed by thereaction of the non-hydrogen-bonded carbonyl group with thiophene.Structural proof of 77 was given by reduction with zinc in aqueousammonia to 2-hydroxy-6-(2-thenyl)-benzoic acid (78), which in turn wasconverted to the corresponding methyl 2-methoxy-6-(2-thenyl)-benzoateand identified by its NOESY spectrum. Ring closure of 78 with zincchloride in glacial acetic acid and acetic anhydride tonaphtho[2,3-b]thiophene 80 proceeded with concomitant acetylation of theoxygen functions. On oxidation with chromium trioxide it afforded thequinone 81, and hydrolysis of the acetoxy function with sodium hydroxidegave the phenolic analogue 82.

Reagents: (a) thiophene, AlCl₃, CH₂Cl₂; (c) Zn, NH₃, Δ; (c) Ac₂O, HOAc,ZnCl₂, Δ; (d) CrO₃, HOAc; (e) 6 N NaOH, Δ.

Compounds of Scheme 2

2-Hydroxy-6-(2-thenoyl)-benzoic acid (77). To a suspension of3-hydroxyphthalic anhydride (1.00 g, 6.09 mmol) and AICl₃ (2.44 g, 18.27mmol) in absolute CH₂Cl₂ (20 mL) and tetramethylethylenediamine (0.15mL, 1.5 mmol) a solution of thiophene (0.49 mL, 6.10 mmol) in absoluteCH₂Cl₂ (10 mL) was added dropwise over 30 min such that the temperatureof the reaction remained below 30° C. The solution was stirred at roomtemperature for an additional 12 h. Then it was treated with ice-water(250 mL), and the product was extracted with CH₂Cl₂ (5×100 mL). Charcoalwas added to the combined organic layer, the mixture was filtered,extracted with 2 N NaOH (3×50 mL) and then acidified with conc. HCl toafford white crystals: 58% yield; mp 168-170° C.; FTIR 3432 (OH), 3151,1681 (CO₂H), 1630 (CO) cm⁻¹; ¹H NMR (DMSO-d₆) δ11.45 (s, br, 1H,exchangeable), 8.40 (dd, 1H, ³J=4.95 Hz, ⁴J=1.21 Hz), 7.52 (dd, 1H,³J=8.35 Hz, ³J=7.43 Hz), 7.39 (dd, 1H, ³J=3.79 Hz, ⁴J=1.21 Hz), 7.21(dd, 1H, ³J=4.95 Hz, ³J=3.79 Hz), 7.13 (dd, 1H, ³J=8.35 Hz, ⁴J=1.09 Hz),6.97 (dd, 1H, ³J =7.43 Hz, ⁴J =1.09 Hz). Anal. (C₁₂H₈O₄S) C,H.

2-Hydroxy-6-(2-thenyl)-benzoic acid (78). To a mixture of zinc dust(3.55 g, 54.3 mmol) and CuSO₄·5 H₂O (0.10 g) in conc. aqueous NH₃ (250mL) was added 77 (1.39 mL 5.60 mmol). The reaction mixture was heated toreflux for 24 h, then filtered while hot, cooled to room temperature,acidified with conc. HCl, and crystallization was completed overnight inan ice-bath to afford white needles: 73% yield; mp 155-158° C.; FTIR3427 (OH), 3290-2620 (CO₂H), 1654 (CO₂H) cm⁻¹; ¹H NMR (DMSO-d₆) δ11.70(s, br, 1H, exchangeable), 10.50 (s, br, exchangeable), 7.29 (dd, 1H,³J=5.13 Hz, ⁴J=1.27 Hz), 7.247.18 (m, 1H), 6.89 (dd, 1H, ³J=5.13 Hz,³J=3.42Hz), 6.816.71 (m, 3H), 4.22 (s, 2H). Anal. (C₁₂H₁₀O₃S) C, H.

4,5-Diacetoxy-naphtho[2,3-b]thiophene (80). A mixture of 78 (0.50 g,2.13 mmol), acetic anhydride (5 mL), glacial acetic acid (12.5 mL), andanhydrous ZnCl₂ (0.20 g, 2.13 mmol) was heated to reflux for 2 h. Thenthe reaction was cooled to room temperature and treated with water (100mL). The product was filtered by suction, dissolved in CH₂Cl₂, purifiedby chromatography and recrystallized from CH₂Cl₂/hexane to afford paleyellow needles: 33% yield; mp 208-209° C.; FTIR 1759 (ester) cm⁻¹; ¹HNMR (CDCl₃) δ8.32 (s, 1H), 7.86-7.11 (m, 5H), 2.50 (s, 3H), 2.44 (s,3H). Anal. (C₁₆H₁₂O₄S) C, H.

5-Acetoxy-naphtho[2,3-b]thiophene-4,9-dione (81) was obtained from 80(0.10 g, 0.33 mmol) as described for 62b. In addition, the mother liquorwas extraced with CH₂Cl₂ (30 mL), the organic layer washed with water(3×50 mL), the product was purified by chromatography usingCH₂Cl₂/hexane (3/1) and recrystallized from CH₂Cl₂hexane to affordbright-yellow needles: 55% yield; mp 210° C.; FTIR 1752 (ester), 1666(CO), 1590 cm⁻¹; ¹H NMR (CDCl₃) δ8.23 (dd, 1H, ³J=7.75 Hz, ⁴J=1.31 Hz),7.76 (dd, 1H, ³J=8.08 Hz, ³J=7.75 Hz), 7.73 (d, 1H, ³J=5.08 Hz), 7.64(d, 1H, ³J=5.08 Hz), 7.39 (dd, 1H, ³J=8.08 Hz, ⁴J=1.31 Hz), 2.49 (s,3H). Anal. (C₁₄H₈O₄S) C, H.

5-Hydroxy-naphtho[2,3-b]thiophene-4,9-dione (82). A solution of 81 (0.03g, 0.11 mmol) in CH₂Cl₂ (10 mL) and 6 N NaOH was heated to reflux for 12h, until the yellow solution turned into deep violet. Then the reactionmixture was poured into ice-water (50 mL), acidified with conc. HCl, andthe aqueous layer was extracted with CH₂Cl₂ (20 mL). The combinedorganic layer was dried over Na₂SO₄, purified by chromatography andrecrystallized from hexane to afford orange crystals: 92% yield; mp199-200° C.; FTIR 3440 (OH), 1656 (CO), 1632 (CO . . . HO) cm⁻¹; ¹H NMR(CDCl₃) δ12.36 (d, 1H,⁵J=0.44 Hz), 7.79 (dd, 1H, ³J=7.47 Hz, ⁴J=1.20Hz), 7.76 (d, 1H, ³J=5.08 Hz), 7.70 (d, 1H, ³J=5.08 Hz), 7.63 (m, 1H,³J=7.47 Hz, ³J=8.42 Hz, ⁵J=0.44 Hz), 7.29 (dd, 1H, ³J=8.42 Hz, ⁴J=1.21Hz). Anal. (C₁₂H₆O₃S) C, H.

Various compounds of formula III are shown in Tables 3 and 4, below.TABLE 3

Compound X R¹ R² 7 O OH COMe

TABLE 4 III

Compound X R¹ R² 45 S H H 81 S 5-OCOMe H 82 S 5-OH H

Example 4. Activity of Synthetic Lapcho Analogs

Compounds of the present invention have demonstrated potentantiproliferative activity against the nontransformed human keratinocyteline HaCaT, as demonstrated by reduction in cell number over time ascompared to control plates. Anthralin, an antipsoriatic drug, was usedas a positive control. Antiproliferative activity was measured directlyby counting the dispersed cells under a phase-contrast microscope.

HaCaT keratinocyte proliferation assay and LDH release were describedpreviously in full detail (Muller et al. J. Med. Chem. 39: 3132-3138,1996; Müller et al. J. Med. Chem. 37: 1660-1669, 1994). For the cancercell line studies, exponentially growing cells were seeded at 1,000cells per well in six-well plates and allowed to attach for 24 h.Compounds of the invention or β-lapachone, solubflized in DMSO, wereadded to the wells in micromolar concentrations. Control wells weretreated with equivalent volumes of DMSO. After 4 h the supernatant wasremoved and fresh medium was added. Cultures were observed daily for10-15 days and then were fixed and stained. Colonies of greater than 30cells were scored as survivors.

Table 5 shows the concentrations of the compounds required to inhibit50% of cell growth (IC₅₀). The cytotoxicity of naphthoquinones has beenthought to result, at least in part, from reactive oxygen species,generated during redox cycling between the quinine and reductionproducts (Munday, R., Free Radic. Biol. Med., 22, 689-695, 1997), whichcause peroxidative damage to membrane lipids. To assess the correlationof keratinocyte growth inhibition with membrane damage, the release oflactate dehydrogenase from the treated cells was also quantitated. TABLE5 AA^(a) in AA^(c) in Cancer Cell Lines HaCaT Cells LDH^(b) IC₅₀ (μM)Compound X R R1 R2 IC₅₀ (μM) (mU) DLD1 SW480 MCF7 Compounds of Formula I73a O Me NA NA >5 ND >32 >32 >32 74a O Ph NA NA 1.9 142 7.6 12 ≦1 75a O3,4-(OMe)₂—Ph NA NA >5 ND 14 30 12 63a S Me NA NA 5.0 ND >32 >32 >32 64aS Ph NA NA 0.3 122 1.5 ≦1 ≦1 62a S 4-OMe—Ph NA NA >5 ND >32 >32 >32 65aS 3,4-(Ome)₂—Ph NA NA >5 ND >32 >32 >32 66a S 4-NO₂—Ph NA NA >5ND >32 >32 >32 Compounds of Formula II  6 O Me NA NA 0.5 331 0.8 ≦1 ≦174b O Ph NA NA 0.7 222 ND ND ND 75b O 3,4-(OMe)₂—Ph NA NA 2.5 250 1.4 NDND 63b S Me NA NA 0.3 134 1 4 ≦1 64b S Ph NA NA 1.7 ND 1.3 4 ≦1 62b S4-OMe—Ph NA NA >5 ND 5.5 16 3 65b S 3,4-(OMe)₂—Ph NA NA 0.8 137 2.6 10≦1 62c S 4-OH—Ph NA NA 2.7 123 5.3 12 2 65c S 3,4-(OH)₂—Ph NA NA 1.5 11811 20 8 66b S 4-NO₂—Ph NA NA 4.0 ND 25 ND ND Compounds of Formula III  7O NA 8-OH COMe 0.3 346 ND ND ND 45 S NA H H >5 222 ND ND ND 81 S NA5-OCOMe H 1.4 160 ND ND ND 82 S NA 5-OH H 1.0 117 ND ND ND α-lapachoneNA NA NA NA 10 ND ND ND ND β-lapachone NA NA NA NA 0.7 329 4 4 ≦1anthralin NA NA NA NA 0.7 294 NA NA NA vehicle NA NA NA NA NA 135 NA NANA^(a)Antiproliferative activity against HaCaT cells. Inhibition of cellgrowth was significantly different with respect to that of the control,N = 3, p < 0.05.^(b)Activity of LDH (mU) release in HaCaT cells after treatment with 2μM test compound, N = 3, SD < 10%, p < 0.05. NA = not applicable.^(c)Antiproliferative activity against colon cancer cell lines DLD1 andSW480 and breast cancer cell line MCF7. ND = not determined. NA = notapplicable.

As shown in Table 5, treatment of HaCaT cells with anthralin waseffective at inhibition of proliferation (IC₅₀=0.7 μM) but causedsubstantial cellular damage, with LDH release significantly higher thanvehicle controls. Similarly, β-lapachone and the 2-acetylatednaphtho[2,3-b]furan4,9-diones (compounds 6 and 7) inhibited cellproliferation but caused significant LDH release as compared to vehicle.However, several of the thiophene analogs (compounds 63a, 64b, 65b, 65c,81 and 82) inhibited cell proliferation at concentrations comparable toβ-lapachone and the furan analogs but without significant elevation ofLDH release over the vehicle control.

Compounds of this invention were also effective at inhibitingproliferation of human cancer cells including cells from the coloncancer lines DLD1 and SW480 and from the breast cancer line MCF7. Asshown in Table 5, IC₅₀ values in the low micromolar range and below wereobtained for several of these compounds in all three cancer cell lines

The antiproliferative activity of the present synthetic lapachoderivative compounds suggests that compounds of the invention may beexpected to show wide anticancer activity. For example, the compounds ofthe invention may be effective for treating cancers such as breastcancer, leukemia, lung cancer, ovarian cancer, brain cancer, livercancer, pancreatic cancer, prostate cancer, and colorectal cancer. Thesetreatments may be accomplished utilizing the present lapacho derivativecompounds (Formula I, II or III) alone or in combination with otherchemotherapy agents or with radiation therapy. In a preferred embodimentthe present lapacho derivative compounds are used for the prevention ortreatment of cancer (e.g., as a preventative drug) by preventing cancercell formation.

As described in part above, a variety of cancer cell lines could be usedto determine the effectiveness of the novel lapacho derivatives of thepresent invention, including SK-OV-3 and OVCAR-3 human ovarian carcinomacells; SW480, HT-29 and HCT-116 human colon carcinoma cells; MCF-7 andMDA-MB-231 human breast carcinoma cells; MIA PACA-2 and BXPC-3 humanpancreatic carcinoma cells; NCI-H226 and A549 human lung carcinomacells; and DU-145 and PC-3 human prostate cancer cells. Sinceβ-lapachone induces apoptosis only in cancer cell lines and not innormal cells (Li., Y, et al., PNAS, (2003), 100(5), 2674-2678) thepresent compounds can also be tested in a panel of normal cell linesincluding NCM 460 normal colonic epithelial cells and MCF 10A normalbreast epithelial cells.

The results of experiments with β-lapachone and similar chemicalcompounds have shown that the present lapacho derivatives may have astrong apoptotic effect on a variety of human cancer cells and that theycan inhibit growth of other human cancer cells.

OTHER EMBODIMENTS

While the invention has been described in conjunction with the detaileddescription thereof, the foregoing description is intended to illustrateand not limit the scope of the invention, which is defined by the scopeof the appended claims. Other aspects, advantages, and modifications arewithin the scope of the following claims.

1. A compound of formula I:

wherein X is O or S; and R is straight-chained or branched C₁-C₆ alkyl,aryl, substituted aryl, or straight-chained or branched alkylaryl, or apharmaceutically acceptable salt thereof; wherein 1) R is not methyl; 2)where X is O, R is not bromomethyl, unsubstituted phenyl, or phenylsubstituted at the 4-position with methyl, chloro, ethenyl, or2′-chloroethyl; and 3) where X is S, R is not 2-carboxyphenyl.
 2. Acompound according to claim 1, wherein X is S, and R is aryl orsubstituted aryl.
 3. A compound according to claim 1 or 2, wherein saidsubstituted aryl is substituted with at least one substituent selectedfrom the group consisting of hydroxyl, alkoxy, C₁-C₆ alkyl, nitro,halogen, carboxyl and carboxyalkyl.
 4. A compound according to claim 1,wherein X is S, and R is phenyl.
 5. A compound of formula II:

wherein X is O or S; and R is straight-chained or branched C₁-C₆ alkyl,aryl, substituted aryl, or straight-chained or branched alkylaryl, or apharmaceutically acceptable salt thereof; wherein R is not methyl.
 6. Acompound according to claim 5, wherein X is O or S, and R is C₁-C₆alkyl, aryl or mono- or di-substituted aryl.
 7. A compound according toclaim 5 or 6, wherein said substituted aryl is substituted with at leastone substituent selected from the group consisting of hydroxyl, alkoxy,alkyl, nitro, halogen, carboxyl and carboxyalkyl.
 8. A compoundaccording to claim 5, wherein X is O, and R is phenyl.
 9. A compoundaccording to claim 5, wherein X is O, and R is 3,4-dimethoxyphenyl. 10.A compound according to claim 5, wherein X is S, and R is phenyl.
 11. Acompound according to claim 5, wherein X is S, and R is3,4-dimethoxyphenyl.
 12. A compound according to claim 5, wherein X isS, and R is 4-hydroxyphenyl.
 13. A compound according to claim 5,wherein X is S, and R is 3,4-dihydroxyphenyl.
 14. A compound of formulaIII:

wherein X is O or S; R₁ is independently at each incidence hydroxyl,alkoxyl, C₁-C₆ alkyl, nitro, halogen, carboxyl or carboxyalkyl; R₂ ishydrogen or —C(O)—R₃, R₃ is straight-chained or branched C₁-C₆ alkyl,aryl, substituted aryl, or straight-chained or branched alkylaryl; and nis 0, 1 or 2; or a pharmaceutically acceptable salt thereof; wherein 1)where X is O, R₂ is not H; 2) where X is O, and R₂ is —C(O)—R₃, and R₃is methyl, then R₁ is not hydroxyl or methoxy; and 3) where X is S andR₂ is H, then n is 1 and R₁ is selected from —OH and—OC(O)-alkyl(C₁-C₆); and 4) where X is S and R₂ is —C(O)—R₃, and R₃ ismethyl, then R₁ does not represent a 7-acetyl group.
 15. A compoundaccording to claim 14, wherein X is S, R₁ is hydroxyl or alkylcarbonyl,R₂ is hydrogen, and n is
 1. 16. A compound according to claim 14,wherein X is S, R₁ is 5-carboxymethyl, R₂ is hydrogen, and n is
 1. 17. Acompound according to claim 14, wherein X is S, R₁ is 5-hydroxyl, R₂ ishydrogen, and n is
 1. 18. A pharmaceutical composition comprising atherapeutically effective amount of a compound according to claim 1 incombination with a pharmaceutically acceptable carrier.
 19. Apharmaceutical composition comprising a therapeutically effective amountof a compound according to claim 4 in combination with apharmaceutically acceptable carrier.
 20. A pharmaceutical compositioncomprising a therapeutically effective amount of a compound according toclaim 5 in combination with a pharmaceutically acceptable carrier.
 21. Apharmaceutical composition comprising a therapeutically effective amountof a compound according to any one of claims 8-13 in combination with apharmaceutically acceptable carrier.
 22. A pharmaceutical compositioncomprising a therapeutically effective amount of a compound according toclaim 14 in combination with a pharmaceutically acceptable carrier. 23.A pharmaceutical composition comprising a therapeutically effectiveamount of a compound according to claims 16 or 17 in combination with apharmaceutically acceptable carrier.
 24. A method of treating orpreventing cell proliferative disorders comprising administering to amammal a therapeutically effective amount of a compound according to anyof claims 1-17.
 25. The method according to claim 24, wherein thecompound is administered as a pharmaceutical composition of claim 18.26. The method according to claim 24, wherein the compound isadministered as a pharmaceutical composition of claim
 19. 27. The methodaccording to claim 24, wherein the compound is administered as apharmaceutical composition of claim
 20. 28. The method according toclaim 24, wherein the compound is administered as a pharmaceuticalcomposition of claim
 21. 29. The method according to claim 24, whereinthe compound is administered as a pharmaceutical composition of claim22.
 30. The method according to claim 24, wherein the compound isadministered as a pharmaceutical composition of claim 23
 31. A method oftreating cancer or precancerous conditions or preventing cancercomprising administering to a mammal in need thereof a therapeuticallyeffective amount of a pharmaceutical composition according to claim 18.32. A method of treating cancer or precancerous conditions or preventingcancer comprising administering to a mammal in need thereof atherapeutically effective amount of a pharmaceutical compositionaccording to claim
 19. 33. A method of treating cancer or precancerousconditions or preventing cancer comprising administering to a mammal inneed thereof a therapeutically effective amount of a pharmaceuticalcomposition according to claim
 20. 34. A method of treating cancer orprecancerous conditions or preventing cancer comprising administering toa mammal in need thereof a therapeutically effective amount of apharmaceutical composition according to claim
 21. 35. A method oftreating cancer or precancerous conditions or preventing cancercomprising administering to a mammal in need thereof a therapeuticallyeffective amount of a pharmaceutical composition according to claim 22.36. A method of treating cancer or precancerous conditions or preventingcancer comprising administering to a mammal in need thereof atherapeutically effective amount of a pharmaceutical compositionaccording to claim
 23. 37. A method of treating or preventing psoriasiscomprising administering to a mammal in need thereof a therapeuticallyeffective amount of a pharmaceutical composition according to claim 18.38. A method of treating or preventing psoriasis comprisingadministering to a mammal in need thereof a therapeutically effectiveamount of a pharmaceutical composition according to claim
 19. 39. Amethod of treating or preventing psoriasis comprising administering to amammal in need thereof a therapeutically effective amount of apharmaceutical composition according to claim
 20. 40. A method oftreating or preventing psoriasis comprising administering to a mammal inneed thereof a therapeutically effective amount of a pharmaceuticalcomposition according to claim
 21. 41. A method of treating orpreventing psoriasis comprising administering to a mammal in needthereof a therapeutically effective amount of a pharmaceuticalcomposition according to claim
 22. 42. A method of treating orpreventing psoriasis comprising administering to a mammal in needthereof a therapeutically effective amount of a pharmaceuticalcomposition according to claim 23.